The overall goal of this proposal is to dissect transcription mechanisms in the human dopamine transporter gene (hDAT). The dopamine transporter (DAT) regulates the spatio-temporal domains of dopamine neurotransmission by reuptake and release of dopamine and thus contributes to locomotion, motivation, cognition and attention, working memory, behavioral organization and hormone release. It is well recognized that expression of the DAT gene in the brain is highly circumscribed, varies among individual subjects and can be regulated by endogenous and exogenous factors such as cocaine abuse. Altered DAT expression may contribute to hDAT-associated pathophysiological states such as substance abuse. However, information about how hDAT expression is regulated and how DNA sequence variation influences the regulated expression remains sporadic. The hypothesis to be tested in this proposal is that transcriptional antagonisms play a major role in regulating hDAT activity. Our preliminary studies show that different cis-acting elements across the entire gene display either enhancing or silencing activities. Therefore, two specific aims of this project are to: 1) demonstrate transcriptional antagonisms between the cis-acting elements and their correlation with hDAT expression; and 2) identify transcription factors that bind to the cis-acting elements around the hDAT promoter in human postmortem brain tissues. The results will add fundamental knowledge on lead sites through which hDAT is regulated by external factors including drugs of abuse and confers risks for related brain disorders.